The envelope glycoproteins of HIV-1 are initially synthesized as a single 160 kD precursor, gp160, which is cleaved at the Arg511-Ala512 bond by a cellular protease, producing gp120 and the integral membrane protein gp41 (Kido, H., et al., J. Biol. Chem. 268:13406-13413 (1993)). The biological activity of gp120 is a key ingredient in initial binding of host cells by HIV-1, propagation of the virus and its toxic effects on uninfected neurons and other cells (Kieber-Emmons, T., et al., Biochim. Biophys. Acta. 989:281-300, (1987); Capon et al., Ann. Rev. Immunol. 9:649-678). Thus, gp120 is a target of passive and active immunization against AIDS (Kahn, J. O., et al., J. Infec. Dis. 170:1288-1291 (1994); Birx, D. L., et al., Curr. Opin. Immunol. 5:600-607 (1993); Berman, P. W., et al., Proc. Natl. Acad. Sci. USA 85:5200-5204 (1988)). Binding of a conformational epitope of gp120 to CD4 receptors on host cells is the first step in HIV-1 infection. Individual amino acids constituting this epitope appear to be located in the second (C2), third (C3), and fourth (C4) conserved gp120 segments (Olshevsky, T. J., et al., J. Virol. 64:5701-5705 (1990)). These are gp120 residues 256, 257, 368-370, 421-427 and 457. Monoclonal antibodies that bind the CD4 binding site have been described (Thali, M., et al., J. Virol. 65:6188-6193 (1991); Thali, M., et al., J. Virol. 66:5635-5641 (1992)). Since the CD4 binding site is a conformational epitope, distant residues that are not themselves constituents of the epitope may be important in maintaining the ability to bind CD4. Gp120 interactions with other host cell proteins are also essential for virus propagation. For example, binding of gp120 by calmodulin may be involved in HIV-1 infectivity, as revealed by the inhibitory effect of calmodulin antagonists (Srinivas, R. V., et al., AIDS Res. Hum. Retroviruses 108:1489-1496 (1994). Asp180 located between the V1 and V2 regions of gp120 is critical for viral replication (Wang, W-K., et al., J. Virol. 69:538-542 (1995)). Similarly, the V3 loop is essential for infectivity (Ivanoff, L. A., et al., Virology 187:423-432 (1992)). It is clear, therefore, that structural determinants in gp120 other than those constituting the CD4 binding site are likely to be necessary for virus genome replication, coat protein synthesis and virus particle packaging.
Shedding of gp120 by virus particles and infected cells has been proposed as a factor in the pathogenesis of AIDS (Gelderblom, H. R. et al., Lancett ii:1016-1017 (1985)). Purified gp120 is toxic for cultured neurons (Brenneman, D. E. et al., Nature 335:639-642 (1988); Muller, W. E. G. et al., Eur. J. Pharmacol 226:209-214 (1992)). Uninfected T-lymphocytes coated with gp120 can be lysed by antibody-dependent monocytes (Hober, D. et al., FEMS Immunol. Med. Microbiol. 10:83-92 (1995)). Uninfected CD4+ cells can die because binding of the gp120-gp41 complex induces apoptosis (Laurent-Crawford, A. G. et al., Res. Virol 146:5-17 995)). Gp120 also binds complement components (Stoiber, H. et al., AIDS 9:19-26 (1995)).
There has been considerable interest in the possibility that gp120 contributes to neural damage observed in AIDS patients (Everall, I. P et al., J. Neuropathol. Exp. Neurol. 52:561-566 (1993)). Productive infection in the brain occurs in a relatively small number of cells, which include microglia, multinucleated giant cells and blood-derived macrophages (Epstein. L. G et al., Ann. Neurol. 33:429-436 (1993)). Yet, there is wide-spread brain damage at areas that are not infected by the virus. This has lead to suggestions that soluble gp120 shed by the virus and cytokines produced by infected cells may be responsible for the damage (Lipton, S. A., Brain Pathol. 1:193-199 (1991), Giulian, D. et al., Science 250:1593-1595 (1990); Benos, D. J. et al., Proc. Natl. Acad. Sci. USA 91:494-498 (1994)). The neurotoxic effects of gp120 may be indirect, involving stimulation of the NMDA receptor (Benos, D. J. et al., Proc. Natl. Acad. USA 91:494-498 (1994)) or induction of cytokines (Yeung, M. C. et al., AIDS 9:137-143, (1995)). Gp120 also stimulates neurotoxin release from monocytes (Giulian, D. et al., Proc. Natl Acad. Sci. USA 90:2769-2773, (1993)), indicating that its neurotoxic effects may require the participation of this cell type. The evidence for neurotoxicity of soluble gp120 includes:
(a) transgenic mice expressing gp120 in the brain show widespread neuronal damage (Toggas, S.M. et al., Nature 367:188-193 (1994)); PA1 (b) pure gp120 at sub-picomolar concentrations kills cultured astrocytes and cerebral cortical cells ((Brenneman, D. E. et al., Nature 335:639-642 (1988); Muller, W. E. G. et al., Eur. J. Pharmacol 226:209-214 (1992)).); PA1 (c) injections of pure gp120 in vivo produce brain damage (Hill, J. M. et al., Brain Res. 603:222-223 (1993)); PA1 (d) a gp120-like neurotoxic activity present in spinal fluid has been described (Buzy, J. et al., Brain Res. 598:10-18 (1992)); PA1 (e) peptide-T, an octapeptide corresponding to a segment of gp120 and bearing some homology to the neuropeptide VIP, may ameliorate neuronal dysfunction in AIDS patients (Bridge, T. P. et al., Psychopharmacol. Bull 27:237-245 (1991)).
gp120 expresses many linear and conformational antigenic epitopes to which antibodies are made. These include neutralizing antibodies made by HIV-infected individuals to the V3 loop (Dreyer, E. B. et al., Science 248:364-367 (1990); Meylan, P. R. et al., AIDS 6:128-130 (1992); Pollard, S. et al., Proc. Natl. Acad. Sci. USA 88:11320-11324 (1991)). Because the V3 loop is a hypervariable region, these antibodies are type-specific, with neutralizing activity directed only against HIV variants with V3 sequences similar to the virus strain responsible for initial infection. On the other hand, antibodies made in the later stages of the disease can be broadly protective, as measured by inhibition of the ability of different HIV-1 strains to infect susceptible cell lines and primary lymphoid cell cultures in vitro. A subset of these protective antibodies are directed against conserved regions of gp120 essential for binding the CD4 receptor and virus propagation (Hattori, T. et al., FEBS Lett 248:48-52 (1989); Schulz, T. et al., AIDS Res. Hum. Retroviruses 9:159-166 (1993); Clements, G. J. et al., AIDS Res. Hum. Retroviruses 7:3-16 (1991)). It is widely believed that recruitment of neutralizing antibody responses to the conserved regions of gp120 will be necessary for effective vaccination against AIDS. Similarly, the success of passive immunization with antibodies will depend on the ability to recognize conserved gp120 regions.
Others have observed that autoantibodies found in systemic lupus erythematosus patients are able to bind gp120 (Gu, R. et al., AIDS Res. Hum. Retroviruses 9:1007-1015 (1993; Callebaut, C, et al., Science 262:2045-2050 (1993)). Cross-reactivity between antibodies to gp120 and to HLA class I heavy chains (H-chains) has been suggested (Sattentau, Q. J. et al., J. Virol. 67:7383-7393 (1993)). AIDS patients have also been described to express DNA-hydrolyzing catalytic antibodies (Woolley, D. W. et al., John Wiley & Sons, Inc. 1952 pp 82).
Speculations that antibodies may develop catalytic activity date back to Woolley in 1952 (Gao, Q. S., et al., J. Biol. Chem 269:32389-32393 (1994)) who suggested that if exposed to an antigen for a sufficiently long period, the immune system may synthesize catalytic antibodies. A detectable sequence homology exists between antibody L-chains and serine proteases (Sun, M. et al., J. Biol. Chem 269:734-738 (1994)). Kohen et al. reported that immunization with steroids or dinitrophenol conjugated to carrier proteins provoked the formation of antibodies with esterase capabilities (Sun, M. et al., J. Immunol 153:5121-5126 (1994); Paul, S. et al., Appl. Biochem. Biotechnol. 47:241-255 (1994)). Human autoantibodies that hydrolyze the neuropeptide VIP have been demonstrated (Li, L., et al., Mol. Immunol. 1996, In press.). Autoantibody catalyzed hydrolysis of VIP has been reproduced (Li, L. et al., J. Immunol. 154:3328-3332 (1995)). Other groups have shown autoantibody mediated hydrolysis of DNA (Tyutyulkova, S. et al., Biochimica Biophysica Acta 1996, In press.; Kalaga, R. et al., J. Immunol 155:2695-2702 (1995)).
Antibodies with enzymatic activity offer the possibility of specific, high efficiency catalytic chemical conversion of ligands. Many biological mediators are peptides or proteins, including the antigens of pathogenic organisms, hormones, neurotransmitters and tumor specific antigens. It is possible to utilize the vast repertoire of specificities that the immune system encompasses to catalyze chemical reactions not within the scope of naturally occurring enzymes. The combination of antibody specificity with the catalytic power of enzymes has the potential of generating potent therapeutic agents, i.e., catalytic antibodies capable of specifically hydrolyzing key viral envelope proteins.